DNA synthesis: The basics

During cell division, a cell must duplicate its DNA so that each cell receives a complete set of genetic instructions. DNA replication involves the synthesis of a complementary DNA strand for each of the original DNA strands. The process follows a semi-conservative model, where each new DNA molecule contains one strand from the original molecule and one newly synthesized strand. DNA is double-stranded and includes sense strands (code in the 5’ to 3’ direction) and antisense strands (the non-coding DNA strand of a gene).

The key steps in DNA synthesis include:

• Initiation: The process begins at specific sites on the DNA molecule called origins of replication. Enzymes called helicases unwind and separate the DNA strands, creating a replication fork.
• Primer synthesis: Primers, short RNA sequences, are synthesized by an enzyme called primase. Primers provide a starting point for DNA synthesis.
• DNA synthesis: DNA polymerase enzymes catalyze the addition of nucleotides to the growing DNA strand. Nucleotides are the building blocks of DNA, and they pair with complementary bases on the template strand (adenine with thymine, and guanine with cytosine). DNA polymerase adds new nucleotides to the 3' end of the growing strand.
• Proofreading: DNA polymerase also has proofreading capabilities to ensure accuracy in base pairing. Incorrectly paired nucleotides are removed and replaced with the correct pair.
• Ligation: After the synthesis of the new DNA strand, the fragments (Okazaki fragments in the lagging strand) are joined together by enzymes called ligases.

DNA synthesis is a highly regulated and an intricate process involving various enzymes and accessory proteins. It plays a crucial role in maintaining the integrity and stability of genetic information in living organisms.

DNA synthesis performed outside of a cell has various important applications across different fields of science and technology. There are two types of synthesis: natural and synthetic. Synthetic DNA synthesis is a technique that allows scientists to create DNA molecules without a template and in almost any sequence.  Some of the key uses of DNA synthesis include:

Research and biotechnology

• Gene cloning: DNA synthesis is used to create copies of specific DNA fragments or genes for further study.
• Site-directed mutagenesis: Researchers can introduce specific mutations into a DNA sequence for functional studies.
• Constructing plasmids and vectors: DNA synthesis is crucial for creating customized plasmids and vectors used in genetic engineering and molecular biology experiments.

Genome editing

• CRISPR-Cas9 technology: Custom DNA synthesis is integral to the design and synthesis of guide RNAs and DNA repair templates for genome editing applications using CRISPR-Cas9 or other genome-editing techniques.

DNA sequencing

• Sequencing controls: Synthetic DNA is used as controls and standards in DNA sequencing experiments to validate the accuracy of the sequencing process. Artificial DNA synthesis permits the identification of novel biological systems. Sequencing is used for confirmation activities.

Diagnostic applications

• Probe synthesis: DNA probes used in various diagnostic techniques, such as fluorescence in situ hybridization (FISH) or polymerase chain reaction (PCR), are often synthesized to specifically target and detect particular DNA sequences.

Vaccine development research

• Antigen synthesis: DNA synthesis is used to create synthetic genes that encode antigens for the development of vaccines. DNA fragments and genes can be used as in vitro transcription templates for mRNA production.

Therapeutic proteins

• Recombinant DNA technology: This involves inserting DNA that encode the protein into bacterial or mammalian cells, expressing the protein in those cells, and then purifying it from them.

Antibodies

• Detection: Antibodies can be used to detect newly synthesized DNA.

Drug discovery research

• Library synthesis: DNA synthesis is employed to generate libraries of DNA sequences, which can be screened for potential drug candidates or used in high-throughput screening assays. This is just one example, but there are many more, including protein engineering and CRISPR library screening.

Synthetic biology

• Designing synthetic genomes: DNA synthesis plays a crucial role in the field of synthetic biology, where researchers design and engineer novel organisms with specific functions or traits.
• Gene writing: Genome engineering technology that writes therapeutic code into a genome in order to treat a disease.

Personalized medicine

• Customized DNA constructs: In personalized medicine, synthetic DNA can be used to create customized therapeutic constructs, such as personalized gene therapies or gene editing tools tailored to individual patients.

Functional genomics

• Synthetic genes for functional studies: DNA synthesis facilitates the creation of synthetic genes for studying their function or regulatory elements in cellular processes.

Academic and educational purposes

• Teaching and training: Synthetic DNA is often used in educational settings to teach molecular biology concepts and techniques.